Polarized epithelial cells are important in regulating the environment between external and internal compartments in an organism. Many malignant and nonmalignant diseases are characterized by changes in cell polarity. It is not known whether these are a cause or effect of the specific disease, but an understanding of the biogenesis, transport, assembly and maintenance of polarized cells may lead to important insights on the mechanisms and/or treatments of these diseases. Studies providing the most insight into these processes have utilized, as models, the infection of polarized monolayers of MDCK cells by enveloped RNA viruses. Such studies have shown that certain viruses such as influenza virus and Sendai virus bud only from the apical domain, whereas other viruses such as Vesicular Stomatitis virus bud exclusively from the basolateral domain. This project proposes to clone, express and follow the transport in eukaryotic cells of proteins encoded in the viral genome of one of these viruses, Sendai virus, and its variant, F1-R. The two viruses differ in their site of budding in polarized cells. Wild-type virus bud exclusively from the apical domain, whereas F1-R buds from both apical and basolateral domains. In other viral systems, the envelope glycoproteins, when expressed without any other viral genes, have been shown to localize in the same domain from which the virus ultimately buds. This indicates that their polypeptide backbones contain a signal for polarize transport and the proteins may be responsible for directing the budding location of the virus. We will first test to see where the Sendai viral proteins are transported. By using individual and coexpression studies, we hope to identify the gene in F1-R that contains the mutation causing the difference in budding pattern. Utilizing recently published information concerning the sequence difference of the viruses and applying it to the gene we have determined to be responsible for the budding differences, we will use site-specific mutagenesis to systematically change variant sequences back to wild-type. By following the transport of the mutagenized protein, we hope to identify the mutation that causes the budding difference in F1-R and to possibly identify an apical transport signal.